Humoral hypercalcemia of malignancy (HHM) is a common and life- threatening complication of certain tumors. Tumor-derived factors are understood to cause hypercalcemia by acting peripherally on bone and kidney but the mechanism by which this occurs and the nature of the factors have been controversial. One theory proposed that circulating tumor derived factors interacted with the parathyroid hormone (PTH) receptors to mimic the biological actions of the hormone. The recent structural identification of a human, tumor-derived hypercalcemic factor (hHCF) revealed a peptide with striking homology to PTH within the first 14 amino acids of the N-terminus. Our preliminary studies, using a synthetic fragment, hHCF-(1-34)NH2, suggest that the factor binds to PTH receptors and stimulates cAMP in vitro. In vivo, it produces hypercalcemia, acts on bone and kidney and stimulates 1,25-dihydroxyvitamin D3 formation with an apparent potency 6-10 fold greater than that of PTH. The main aims of the proposed work are to characterize the biological activity of various derivatives of the hHCF using unique in vivo and in vitro bioassay systems. Our first series of experiments will attempt to delineate the mechanism by which the factor produces hypercalcemia with particular emphasis on the role of 1,25- dihydroxy vitamin D3 and bone resorption to this process. A second level of experimentation will seek to establish the structural features of the hHCF that account for its enhanced potency and to determine whether synthetic PTH antagonists are useful in blocking its action. Such information may eventually lead to the design of antagonists for treatment of the disease. Collectively, we believe our studies will provide critical information on the pathogenesis of human hypercalcemia of malignancy and will foster insight into PTH structure-activity relationships for future design of potent hormone analogs.